PROJECT SUMMARY Autophagy is a crucial catabolic pathway by which cellular waste is recycled. Autophagic dysfunction has been implicated in cellular quality control, responses to stress, development, lifespan, and a range of infectious and other diseases in humans, including cancer, neurodegenerative diseases, and diabetes. Membrane fusion is a critical process involved in autophagosome biogenesis. The exact molecular mechanism of autophagic membrane fusion remains far from clear and thus a major topic of investigation. Since membrane fusion activity could act as a switch to spatially and temporally regulate the autophagic flux in human diseases due to its dysregulation, dissecting the fusion machinery is essential to understanding the exact roles of autophagy in specific disease contexts. Therefore, studying the regulatory mechanism of membrane fusion will provide the opportunity to develop new therapeutic strategies in order to control activity of autophagy. Based on our preliminary findings, we hypothesize that the morphology and modification status of autophagic SNAREs are important for mediating membrane fusion involved in autophagosome biogenesis, while this process is regulated by accessory proteins including nuclear receptor binding factor 2 and Atg9. Further systematical studies on the role of SNARE-mediated membrane fusion in autophagosome maturation and initiation are critical to elucidate detailed molecular mechanisms, which could offer therapeutic advances. Moreover, an attempt to find new fusogens and reconstitute the early autophagosome biogenesis is an important expansion, which is also essential for future drug development.